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- EMDB-21666: Cryo-EM map of the mutant OCCM (ORC-Cdc6-Cdt1-Mcm2-7 with Mcm6 WH... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-21666 | ||||||||||||||||||
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Title | Cryo-EM map of the mutant OCCM (ORC-Cdc6-Cdt1-Mcm2-7 with Mcm6 WHD truncation) loaded on DNA at 10.5 A resolution | ||||||||||||||||||
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![]() | DNA replication / OCCM-deltaC6 / REPLICATION-DNA complex | ||||||||||||||||||
Function / homology | ![]() CDC6 association with the ORC:origin complex / Cul8-RING ubiquitin ligase complex / maintenance of rDNA / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state / MCM complex binding / nuclear DNA replication / Assembly of the ORC complex at the origin of replication / premeiotic DNA replication ...CDC6 association with the ORC:origin complex / Cul8-RING ubiquitin ligase complex / maintenance of rDNA / MCM core complex / Assembly of the pre-replicative complex / Switching of origins to a post-replicative state / MCM complex binding / nuclear DNA replication / Assembly of the ORC complex at the origin of replication / premeiotic DNA replication / nuclear origin of replication recognition complex / pre-replicative complex assembly involved in nuclear cell cycle DNA replication / mitotic DNA replication / Activation of the pre-replicative complex / CMG complex / single-stranded 3'-5' DNA helicase activity / nuclear pre-replicative complex / MCM complex / Activation of ATR in response to replication stress / DNA replication preinitiation complex / replication fork protection complex / mitotic DNA replication checkpoint signaling / mitotic DNA replication initiation / double-strand break repair via break-induced replication / silent mating-type cassette heterochromatin formation / regulation of DNA-templated DNA replication initiation / single-stranded DNA helicase activity / DNA strand elongation involved in DNA replication / CDK-mediated phosphorylation and removal of Cdc6 / Orc1 removal from chromatin / DNA unwinding involved in DNA replication / regulation of DNA replication / nuclear replication fork / 3'-5' DNA helicase activity / DNA replication origin binding / subtelomeric heterochromatin formation / DNA replication initiation / nucleosome binding / heterochromatin formation / helicase activity / G1/S transition of mitotic cell cycle / chromosome / single-stranded DNA binding / DNA helicase / chromosome, telomeric region / cell division / GTPase activity / DNA damage response / chromatin binding / GTP binding / ATP hydrolysis activity / nucleoplasm / ATP binding / nucleus / metal ion binding / cytoplasm Similarity search - Function | ||||||||||||||||||
Biological species | ![]() ![]() | ||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 10.0 Å | ||||||||||||||||||
![]() | Yuan Z / Schneider S | ||||||||||||||||||
Funding support | ![]() ![]()
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![]() | ![]() Title: Structural mechanism of helicase loading onto replication origin DNA by ORC-Cdc6. Authors: Zuanning Yuan / Sarah Schneider / Thomas Dodd / Alberto Riera / Lin Bai / Chunli Yan / Indiana Magdalou / Ivaylo Ivanov / Bruce Stillman / Huilin Li / Christian Speck / ![]() ![]() Abstract: DNA replication origins serve as sites of replicative helicase loading. In all eukaryotes, the six-subunit origin recognition complex (Orc1-6; ORC) recognizes the replication origin. During late M- ...DNA replication origins serve as sites of replicative helicase loading. In all eukaryotes, the six-subunit origin recognition complex (Orc1-6; ORC) recognizes the replication origin. During late M-phase of the cell-cycle, Cdc6 binds to ORC and the ORC-Cdc6 complex loads in a multistep reaction and, with the help of Cdt1, the core Mcm2-7 helicase onto DNA. A key intermediate is the ORC-Cdc6-Cdt1-Mcm2-7 (OCCM) complex in which DNA has been already inserted into the central channel of Mcm2-7. Until now, it has been unclear how the origin DNA is guided by ORC-Cdc6 and inserted into the Mcm2-7 hexamer. Here, we truncated the C-terminal winged-helix-domain (WHD) of Mcm6 to slow down the loading reaction, thereby capturing two loading intermediates prior to DNA insertion in budding yeast. In "semi-attached OCCM," the Mcm3 and Mcm7 WHDs latch onto ORC-Cdc6 while the main body of the Mcm2-7 hexamer is not connected. In "pre-insertion OCCM," the main body of Mcm2-7 docks onto ORC-Cdc6, and the origin DNA is bent and positioned adjacent to the open DNA entry gate, poised for insertion, at the Mcm2-Mcm5 interface. We used molecular simulations to reveal the dynamic transition from preloading conformers to the loaded conformers in which the loading of Mcm2-7 on DNA is complete and the DNA entry gate is fully closed. Our work provides multiple molecular insights into a key event of eukaryotic DNA replication. | ||||||||||||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 54.2 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 37.6 KB 37.6 KB | Display Display | ![]() |
Images | ![]() | 149.6 KB | ||
Filedesc metadata | ![]() | 13 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 372.3 KB | Display | ![]() |
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Full document | ![]() | 371.8 KB | Display | |
Data in XML | ![]() | 6.2 KB | Display | |
Data in CIF | ![]() | 7 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6wgiMC ![]() 6wgcC ![]() 6wgfC ![]() 6wggC C: citing same article ( M: atomic model generated by this map |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
File | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Voxel size | X=Y=Z: 1.31 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
+Entire : ORC-Cdc6-Mcm-Cdt1-dsDNA
+Supramolecule #1: ORC-Cdc6-Mcm-Cdt1-dsDNA
+Macromolecule #1: DNA (34-MER)
+Macromolecule #2: DNA (34-MER)
+Macromolecule #3: Cell division control protein 6
+Macromolecule #4: Origin recognition complex subunit 1
+Macromolecule #5: Origin recognition complex subunit 2
+Macromolecule #6: Origin recognition complex subunit 3
+Macromolecule #7: Origin recognition complex subunit 5
+Macromolecule #8: Origin recognition complex subunit 4
+Macromolecule #9: Origin recognition complex subunit 6
+Macromolecule #10: DNA replication licensing factor MCM2
+Macromolecule #11: DNA replication licensing factor MCM3
+Macromolecule #12: DNA replication licensing factor MCM4
+Macromolecule #13: Minichromosome maintenance protein 5
+Macromolecule #14: Cell division cycle protein CDT1
+Macromolecule #15: DNA replication licensing factor MCM6
+Macromolecule #16: DNA replication licensing factor MCM7
+Macromolecule #17: PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER
+Macromolecule #18: MAGNESIUM ION
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 7.5 |
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Grid | Details: unspecified |
Vitrification | Cryogen name: ETHANE |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 50.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
Startup model | Type of model: PDB ENTRY |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 10.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 27830 |
Initial angle assignment | Type: PROJECTION MATCHING |
Final angle assignment | Type: PROJECTION MATCHING |
-Atomic model buiding 1
Refinement | Protocol: RIGID BODY FIT |
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Output model | ![]() PDB-6wgi: |